[Phenylhexyl isothiocyanate reducing U266 cell line methylation level of p16 gene].
This study was purposed to investigate whether phenylhexyl isothiocyanate (PHI) can reduce p16 gene methylation level or not. The myeloma U226 cell line was cultured with PHI of 0, 5, 10 micromol/L for 72 hours, then DNA was extracted. Hydrosulfite was used to treat the genome DNA of healthy adult, PCR amplification was carried out by using this DNA as template. The gene chip detecting methylation changes of 3 CpG in promoter region of p16 gene was constructed by designing a pair of probes which contain one methylated and one unmethylated probes. This pair of probes was used to detect 3 consecutive sites of CpG island in p16 gene. The standard curve was constructed by using gene chip after the methylated and unmethylated DNA were mixed at different ratio. Then treated samples of U266 cells were dotted on gene chip, obtained results were compared with standard curve to get the quantitative results. The results indicated that the probes on chip had excellent reproducible ability and precision, the methylation level of p16 gene in U266 cells treated with 0, 5 and 10 micromol/L of PHI was determined as 78.2%, 61.7% and 54.8%, respectively. It is concluded that the PHI can reduce the methylation level of p16 gene in U266 cells.
Chen
Bao-An
BA
Department of Hematology, Zhongda Hospital, Clinical Medical College of Southeast University, Nanjing 210009, Jiangsu Province, China.
Shou
Bei-Ming
BM
Zhou
Dong-Rui
DR
Ding
Jia-Hua
JH
Gao
Chong
C
Sun
Yun-Yu
YY
Wang
Jun
J
Chen
Jian
J
Zhao
Gang
G
Song
Hui-Hui
HH
Bao
Wen
W
Liu
De-Long
DL
Ma
Xu-Dong
XD
Lu
Zu-Hong
ZH
D045744:Cell Line, Tumor; D018899:CpG Islands; D019175:DNA Methylation; D015536:Down-Regulation; D019942:Genes, p16; D006801:Humans; D017879:Isothiocyanates / Q000494:pharmacology*; D020411:Oligonucleotide Array Sequence Analysis
D017879:Isothiocyanates; C073279:6-phenylhexyl isothiocyanate